Yersinia pestis is a Category A pathogen and the causative agent of pneumonic plague. A type III secretion system (T3SS) that is essential for virulence is used by Y. pestis to export a set of proteins known as Yops. Yop effectors (e.g. YopE) are delivered into host cells to disrupt intracellular signaling pathways. Yop effectors are delivered across the host cell plasma membrane by the translocators YopD and YopB. The LcrV protein, localized to the tip of the T3SS, is required to insert YopD and YopB into the plasma membrane and is a well-characterized protective antigen. Y. pestis also assembles protein fibers on its surface that are composed of the F1 protein. F1 is a protective antigen and is being developed together with LcrV as a subunit vaccine. Murine IgG MAbs specific for LcrV or F1 have been shown to confer passive protection against plague in mice. However, it is possible to genetically modify Y. pestis to make it resistant to LcrVand F1-based vaccines or immunotherapeutics, because F1- mutants remain fully virulent, and protective epitopes within LcrV can be altered. Therefore, additional protective antigens need to be identified and developed into targets for vaccines or immunotherapeutics to counteract the threat of genetically modified Y. pestis. This research will identify and characterize protective IgG MAbs specific for YopD and YopB that could be used to passively immunize humans against genetically modified plague. In Aim 1, we will demonstrate passive protection of mice against F1- Y. pestis with MAbs specific for Yop translocator proteins. These experiments will establish a proof-of-principle that pneumonic plague caused by genetically modified Y. pestis can be prevented by MAb immunotherapy. In Aim 2, we will identify epitopes in Yop translocators proteins recognized by protective MAbs. These studies will provide information on the nature of the protective epitopes, and allow for development of a MAb cocktail as an optimized immunotherapy. In Aim 3, we will determine role of constant heavy (CH) regions in protective MAb function. F(ab')2 fragments of protective anti-YopB and anti-YopD MAbs will be generated and tested for protective activity. Chimeric (ch) MAbs containing variable regions of protective MAbs and human CH regions of different isotypes will be constructed, characterized for affinity and specificity, and assayed for protective function. Mice expressing human Fc receptor will be used to measure protective activity of murine-human ch MAbs. Results will increase the usefulness of these MAbs as immunotherapeutics in humans, as well as provide knowledge on the role of constant region in MAb function.